The modification of surfaces of materials to impact specific physical, chemical or biological properties is important in a wide range of bioanalytical settings. The development of strategies that are simple, reproducible and that give stable interfaces are particularly attractive. Immobilization via a photoactive crosslinker is a practically simple and effective method for the covalent attachment of polymers and biomolecules on surfaces. This proposal seeks to investigate the surface chemistry that influences the covalent immobilization of thin films by way of perfluorophenyl azides (PFPAs). The hypothesis is that the composition, structure, and property of the functional surface directly affect the film immobilization yield, efficiency and integrity. We base the hypothesis on our observations that 1) poly (ethylene glycol) and isotactic polypropylene could not be immobilized using our standard spin coating and photoactivation procedure, although the method should in principle be versatile and applicable to any molecules possessing C-H bond; 2) when we varied the density of the surface azido groups by co-depositing a non-photoactive molecule together with the PFPA, the immobilization yield was affected. These observations demonstrate the need for systematic studies on surface and interface properties for the immobilization.chemistry. Our goal is to develop strategies and optimal conditions for the immobilization of a greater variety of molecules and materials, thus making this method truly versatile. Method development includes the fabrication of nanometer-size patterned polymer films and nanowells, and the generation of polymer films and multilayers possessing generic functional groups for bioconjugation. The specific aims are to: 1. Investigate parameters and conditions that influence the immobilization yield, efficiency and integrity of immobilized films. A non-photoactive molecule will be co-adsorbed with the photoactive PFPA to control the density and topography of azido groups on the surface. The functional group on the non-photoactive molecule will be chosen to specifically to enhance the interactions and compatibility with the molecules to be immobilized. 2. Create nanometer-size features of polymer thin films and nanowell arrays using near-field optical lithography. Near-field optical lithography uses an optical probe to directly write features with nanoscale dimensions. In addition, the coupled laser light generates heat at the probe tip. This should increase the yield of immobilization, and broaden the range of materials that can be used for creating nanometer-size structures. We will also use the technique to fabricate true nanowell arrays with nanometer dimensions both spatially and topographically. 3. Generate thin films and multilayers possessing generic functional groups for bioconjugation. This will be accomplished using the direct immobilization chemistry developed in our laboratory. Thin films of polymers that possess -COOH, -NH2 and -SO3H groups will be generated. These polymer films will be used as generic coatings for the conjugation of a variety of biomolecules and other functional materials. [unreadable] [unreadable]